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Blood, Vol. 95 No. 1 (January 1), 2000:
pp. 286-293
IMMUNOBIOLOGY
From the First Department of Internal Medicine, Ehime University
School of Medicine, Shigenobu, Ehime, Japan.
The Wilms tumor (WT1) gene has been reported to be
preferentially expressed in acute leukemia cells, regardless of
leukemia subtype and chronic myelogenous leukemia cells in blast
crisis, but not in normal cells. This finding suggests strongly that
WT1 protein is a potential target of immunotherapy for human leukemia. In this study, we established a CD8+ cytotoxic
T-lymphocyte (CTL) clone directed against a WT1-derived peptide and
examined its immunologic actions on leukemia cells. A
CD8+ CTL clone, designated TAK-1, which lysed
autologous cells loaded with a WT1-derived 9-mer peptide
consisting of the HLA-A24 (HLA-A*2402)-binding motifs was established
by stimulating CD8+ T lymphocytes from a healthy
individual repeatedly with WT1 peptide-pulsed autologous dendritic
cells. TAK-1 was cytotoxic to HLA-A24-positive leukemia cells
expressing WT1, but not to HLA-A24-positive lymphoma cells that did
not express WT1, HLA-A24-negative leukemia cells, or HLA-A24-positive
normal cells. Treating leukemia cells with an antisense oligonucleotide
complementary to the WT1 gene resulted in reduced
TAK-1-mediated cytotoxicity, suggesting that target antigen of TAK-1 on
leukemia cells is the naturally processed WT1 peptide in the context of
HLA-A24. TAK-1 did not inhibit colony formation by normal bone marrow
cells of HLA-A24-positive individuals. Because WT1 is overexpressed
ubiquitously in various types of leukemia cells, but not in normal
cells, immunotherapy using WT1 peptide-specific CTL clones should be an
efficacious treatment for human leukemia. (Blood. 2000;95:286-293)
Cytotoxic T lymphocytes (CTLs) undoubtedly play an
important role in resistance to cancers, including various types of
leukemia, and adoptive transfer of CTLs, which discriminate malignant
and normal cells, should be an efficacious treatment for human
leukemia.1 To develop effective immunotherapy, the specific
tumor antigens that are recognized by the immune system must be
identified. So far, various molecules have been proposed as candidates
for the targets of leukemia-specific CTLs. Among them, fusion proteins resulting from chromosomal translocations are potential targets of
immune responses, as fusion proteins are produced only by leukemia cells, not by normal cells. On the basis of this concept, generation of
fusion protein-specific T lymphocytes has been attempted, and several
investigators have succeeded in establishing T-lymphocyte clones and
bulk cell lines that specifically recognize the synthetic peptide
spanning the fusion point between 2 proteins, such as BCR-ABL in
chronic myelogenous leukemia (CML),2-10 PML-RAR Another strategy for inducing tumor-specific T-lymphocyte responses is
the use of synthetic peptides derived from normal proteins that are
preferentially expressed or overexpressed in tumor cells. Although
various proteins, such as tyrosinase, Pmel/gp100, Melan-A/Mart-1, HER-2/neu, p53, and PRAME, have been identified as the targets of
melanoma- and solid tumor-specific CTLs, only proteinase 3 has been
reported to be a potential target antigen of CTLs directed against
human leukemia cells.14 In this study, we selected Wilms tumor protein (WT1) as a target of CTLs for the development of effective immunotherapy for leukemia, because WT1 recently was found to
be overexpressed in a variety of leukemia cells but not in normal
cells.15-17
The WT1 gene encodes a zinc finger transcription
factor,18 and WT1 binds the early growth response-1 DNA
consensus sequence present in growth factor gene promoters, such as
platelet-derived growth factor A chain, colony-stimulating factor-1,
transforming growth factor- Recently, it was reported that most human leukemia cells aberrantly
overexpress WT1 regardless of the leukemia subtype, and WT1 expression
was proposed to be a leukemia cell marker useful for the diagnosis of
minimal residual disease.16,27,28 The evidence that WT1 is
preferentially expressed in a variety of human leukemia cells, but not
in normal cells, suggests strongly that it may be possible to develop
efficacious immunotherapy for patients with leukemia by targeting this
protein. Therefore, our aim was to generate WT1-specific CTLs and
examine their immunologic actions on leukemia cells. In this study, we
succeeded in establishing a CD8+ CTL clone that recognized
a 9-mer peptide derived from WT1 in the context of HLA-A24, the most
common HLA class I type in all people of Japanese descent (over 60%
are positive). The WT1 peptide-specific CTL clone efficiently lysed
HLA-matched leukemia cells but not HLA-mismatched leukemia cells,
HLA-matched lymphoma cells that did not express WT1, or HLA-matched
normal cells. Furthermore, this CTL clone did not inhibit colony
formation by HLA-matched normal bone marrow cells. In light of these
findings, the possibility of developing immunotherapy for leukemia
using WT1-specific CTLs is discussed.
Cell lines
Synthetic peptide
HLA typing HLA serotyping was performed using a microlymphocyte cytotoxicity test with local qualified antisera. According to the serologic typing results, HLA class I alleles were amplified by the polymerase chain reaction (PCR) using group-specific primers and were then typed at the nucleotide sequence level. In brief, for example, HLA-A9 group alleles, including HLA-A*2402, were amplified using HLA-A9-specific primers and then analyzed using 9 probes that can be used to distinguish 6 alleles (A*2301, A*2402-A*2406), as described previously.32 HLA-A24 expression on some leukemia and lymphoma cells was examined by flow cytometry using a fluorescein isothiocyanate (FITC)-conjugated anti-HLA-A24 monoclonal antibody (MoAb) (One Lambda, Canoga Park, CA) and FITC-conjugated mouse immunoglobulin (Ig) G as the control. HLA-A24 nucleotide sequencing was performed using the dideoxy-chain termination method, as described previously.33Generation of WT1 peptide-specific CTL clones Peripheral blood dendritic cells (DCs) were generated as follows. Monocyte-enriched peripheral blood mononuclear cell (PBMC) fractions were isolated, using a plastic adherence technique, from total PBMCs of HLA-A24-positive healthy individuals. The plastic-adherent cells were cultured further in RPMI 1640 medium supplemented with 10% FCS, 500 U/mL recombinant human interleukin (IL)-4 (Genzyme, Boston, MA), and 800 U/mL recombinant human granulocyte-macrophage colony-stimulating factor (GM-CSF) (Kirin Brewery, Tokyo, Japan). On day 3 of incubation, half of the medium was exchanged for fresh culture medium supplemented with IL-4 and GM-CSF, and culture was continued. On day 5, half of the medium was exchanged for culture medium supplemented with IL-4, GM-CSF, and 100 U/mL recombinant human tumor necrosis factor (TNF)-
(Dainippon Pharmaceutical, Osaka, Japan). On day 8 or 9, the cells were
harvested and used as monocyte-derived DCs for antigen stimulation. The
generated cells appeared to express DC-associated antigens, such as
CD1a, CD80, CD83, CD86, and HLA class I and class II, on their cell surfaces (data not shown). CD8+ T lymphocytes were
isolated, using magnetizable polystyrene beads coated with an anti-CD8
MoAb (DYNAL, Oslo, Norway), from the same donors. A total of 1 million
CD8+ T lymphocytes were cultured with
1 × 105 autologous DCs treated with mitomycin C
(MMC) (Kyowa Hakko, Tokyo, Japan) in RPMI 1640 medium supplemented with
10% heat-inactivated human AB type serum and 5 ng/mL recombinant human
IL-7 (Genzyme) together with a WT1 synthetic peptide at a concentration
of 10 µg/mL in a 16-mm well. After culture for 7 days, half of the
medium was exchanged for fresh culture medium supplemented with IL-7, and the cells were stimulated again by adding
1 × 105 autologous DCs treated with MMC and WT1
peptide at a concentration of 10 µg/mL. After culture for a further 7 days, the cells were stimulated a third time, as they were
for the second except for the addition of IL-7. After
culture for a further 4 days (day 18 of culture), 10 U/mL recombinant
human IL-2 (Boehringer Mannheim, Mannheim, Germany) was added to each
well. The cytotoxicity of the growing cells was then examined, and the
bulk of the cells that were cytotoxic to WT1 peptide-loaded autologous
LCL was cloned by a limiting dilution method, as described
previously.34 T-lymphocyte clones were cultured
continuously in IL-2-containing culture medium, and
MMC-treated autologous PBMCs and the required WT1 peptide were added to
the wells every 2 weeks.
Cytotoxicity assays Chromium-51 release assays were performed as described previously.35 Briefly, 1 × 104 51Cr (Na251CrO4) (New England Nuclear, Boston, MA)-labeled target cells suspended in 100 µL RPMI 1640 medium supplemented with 10% FCS (assay medium) were seeded into round-bottomed microtiter wells and incubated with or without synthetic peptide for 2 hours. In some experiments, the target cells were preincubated with an anti-HLA class I MoAb, w6/32 (ATCC, Rockville, MD), at an optimal concentration (10 µg/mL) for 30 minutes to determine whether cytotoxicity was restricted by HLA class I. Various numbers of effector cells suspended in 100 µL assay medium were added to the well, incubated for 4 hours, and 100 µL supernatant was collected from each well. The percentage of specific lysis was calculated as follows: (cpm experimental release cpm
spontaneous release)/(cpm maximal release cpm spontaneous
release) × 100.
Northern blot analysis Total RNAs were extracted from leukemia and lymphoma cell lines and freshly isolated leukemia cells; 20 µg of total RNAs were separated by electrophoresis on a 1% agarose gel containing 2.2 mol/L formaldehyde and transferred to nylon filters, which were hybridized with 32P ([32P]dCTP;) ICN Radiochemicals, Irvine, CA)-labeled WT1 or glyceraldehyde-3-phosphate dehydrogenase (GAPDH) probes. The WT1 probe used was the 1.4-kb DNA fragment derived from K562 cell line by the reverse transcription-PCR using 5'-CCGAATTCAGGATGTGCGACGTGTGCCT-3' and 5'-CCGAATTCACCTGTATGAGTCCTGGTGTG-3' primers. The extent of hybridization was quantified by scanning the autoradiographs using a Bio-Imaging Analyzer system (BAS1000; Fujifilm, Tokyo, Japan).Treatment of leukemia cells with a WT1 antisense oligonucleotide The 18-base antisense oligonucleotide phosphorothioate complementary to the sequence of the translation initiation site of the WT1 gene was synthesized and purified by high-performance liquid chromatography, as described previously.36 Sense and random sequences of the same 18-base oligonucleotide were also synthesized and used as controls. The oligonucleotide sequences were as follows: sense, 5'-CAGCAAATGGGCTCCGAC-3'; antisense, 5'-GTCGGAGCCCATTTGCTG-3'. Random oligonucleotides were prepared by mixing 4 different 18-base oligonucleotides, heating the mixture at 80°C for 5 minutes, and allowing it to cool slowly to 37°C over 5 hours. The cells were suspended in FCS-free RPMI 1640 medium and placed in a 24-well culture plate. The oligonucleotides were added to the medium at a concentration of 20 µM and incubated at 37°C. After 2 hours, FCS was added to the culture medium at a final concentration of 10%. The same oligonucleotides were added to each well every 24 hours at half the dose of the initial concentration. Three days later, the cells were collected, and their expression levels of WT1 protein were determined by Western blot analysis. These oligonucleotide-treated cells were used as targets in the cytotoxicity assays.Western blot analysis Western blot analysis was performed according to the ECL protocol (Amersham Pharmacia Biotech, Little Chalfont, England, UK). The cells were washed twice with phosphate-buffered saline and then lysed with buffer containing 1% Triton X-100, 100 mg/mL phenylmethylsulfonyl fluoride, 2 mg/mL leupeptin, and 2 mg/mL aprotinin as protease inhibitors. Equal amounts (30 µg) of protein lysate were separated by 10% sodium dodecyl sulfate-polyacrylamide gel electrophoresis and blotted onto Hybond ECL membranes (Amersham Pharmacia Biotech) using the Trans-Biot electrophoretic transfer system (Bio-Rad, Hercules, CA). The protein blots were incubated with a rabbit anti-human WT1 polyclonal antibody (Ab) (WT 180; Santa Cruz Biotechnology, Santa Cruz, CA) or a rabbit anti-human GAPDH polyclonal Ab (Trevigen, Gaithersburg, MD) for 1 hour at room temperature. After washing, each filter was incubated with horseradish peroxidase-labeled goat anti-rabbit IgG and subjected to the enhanced chemiluminescence assay using the ECL detection system (Amersham Pharmacia Biotech).Colony-forming assay The effect of the T-lymphocyte clone on the growth of normal bone marrow cells was examined by performing the colony formation assay described previously8 with a slight modification. Bone marrow cells were isolated from patients with malignant lymphoma without bone marrow invasion and those with nonhematologic disorders undergoing bone marrow examination, after obtaining their informed consent, and cryopreserved until required for use. Cloned T lymphocytes and bone marrow cells, which had been incubated in the presence or absence of WT1 peptide at a concentration of 10 µg/mL for 2 hours were suspended in assay medium at an effector cell to target cell ratio of 3:1, centrifuged at 1000 revolutions per minute for 3 minutes to ensure close cell contact, and then coincubated in assay medium at 37°C for 4 hours. Control bone marrow cells were centrifuged and incubated without the T-lymphocyte clone in the same manner. After incubation, 5 mL of Iscove's modified Dulbecco's medium containing 1% methylcellulose, 5% GCT-conditioned medium, 1% bovine serum albumin, 30% FCS, 100 µM 2-mercaptoethanol, and 3 U/mL erythropoietin (Stem Cell CFU Kit; Baxter, Deerfield, IL) was added to the cell pellet at the final bone marrow cell concentration of 1 × 105 cells/mL. Each cell suspension was then placed in triplicate 24-mm wells and cultured at 37°C for 12 to 14 days, after which, the numbers of colony-forming unit granulocyte-macrophage (CFU-GM) and burst-forming unit erythroid (BFU-E) were counted using an inverted microscope. The significance of differences between values for 2 groups was determined using the paired 2-tailed Student t test, and those at P < .05 were considered significant.
Generation of a WT1 peptide-specific CTL clone We attempted to generate WT1-specific CTLs from 3 healthy HLA-A24-positive individuals using WT1 peptide-pulsed DCs, as described above, and established 1 CTL clone, designated TAK-1, which lysed a WT1 peptide-loaded, but not unloaded, autologous LCL. Flow cytometric analysis demonstrated that more than 99% of TAK-1 cells were CD3+, CD4-, CD8+, and CD56-. The antigen specificity and HLA restriction of TAK-1-mediated cytotoxicity are summarized in Table 1. TAK-1 lysed autologous LCL that was loaded with the WT1-T2 peptide, but was not cytotoxic to unloaded, WT1-T1-, WT1-T3-, or WT1-T4-loaded autologous LCL. TAK-1 also did not lyse autologous LCL loaded with an MAGE-3-derived or an HIV-1 gp41-derived peptide consisting of HLA-A24-high binding affinity. The restriction element of TAK-1 seemed to be HLA-A24, because only HLA-A24-positive allogeneic LCLs were lysed by TAK-1 and TAK-1-mediated cytotoxicity was inhibited by adding the anti-HLA class I MoAb to target cells (data not shown). To confirm HLA-A24 restriction of TAK-1, we examined the cytotoxic activity of TAK-1 to HLA-A*2402 transfectant cell line C1R-A*2402. In the presence of WT1-T2 peptide, TAK-1 was cytotoxic to C1R-A*2402 but not to its parent cell line, C1R. These data show that TAK-1-mediated cytotoxicity was WT1-T2 peptide-specific and restricted by HLA-A24. The peptide titration assay demonstrated that WT1-T2 peptide-specific cytotoxicity was detectable with final peptide concentrations of 1 ng/mL to 100 µg/mL, and the optimal peptide concentration range for TAK-1-mediated cytotoxicity was 1 to 10 µg/mL (Figure 1).
Expression of WT1 mRNA in leukemia and lymphoma cell lines Previous studies demonstrated that WT1 is expressed in most acute leukemia cells and CML cells at blast crisis regardless of the leukemia subtype. First, to determine whether TAK-1 lysed leukemia cell lines in a WT1-specific manner, we carried out Northern blot analysis to evaluate the WT1 messenger RNA (mRNA) expression levels of various leukemia and lymphoma cell lines. Representative data are shown in Figure 2. All the cell lines established from AML, ALL, and CML at blast crisis examined appeared to express abundant WT1 mRNA. In contrast, all the lymphoma cell lines examined expressed no WT1 mRNA. These data affirm those reported previously.
Lysis of leukemia cell lines by TAK-1 We next examined whether TAK-1 was cytotoxic to leukemia cell lines that expressed abundant WT1. As shown in Table 2, TAK-1 showed cytotoxicity to HLA-A24-positive leukemia cell lines despite the absence of added WT1-T2 peptide, whereas no cytotoxicity to HLA-A24-negative leukemia cell lines was detected. Furthermore, TAK-1 appeared to have no cytotoxic activity to lymphoma cell lines, which did not express WT1, regardless of their HLA-A24 expression status. These data suggest strongly that WT1 peptide is processed naturally in leukemia cells, expressed in the context of HLA-A24, and recognized by CD8+ CTLs.
Lysis of freshly isolated leukemia cells by TAK-1 The cytotoxicity of TAK-1 to various kinds of leukemia cells, which were freshly isolated from patients, is shown in Table 3. As observed with the cell lines, TAK-1 was cytotoxic to leukemia cells isolated from HLA-A24-positive patients with AML and ALL, except for some cases in which WT1 may not have been processed efficiently enough to be presented to the T lymphocytes and/or certain molecules important to CTL-mediated cytotoxicity may not have been expressed. Leukemia cells isolated from HLA-A24-negative patients were not lysed by TAK-1, which was not cytotoxic to normal foreskin fibroblasts or PBMCs isolated from HLA-A24-positive or HLA-A24-negative donors. Therefore, TAK-1 appeared to discriminate freshly isolated leukemia cells, as well as leukemia cell lines, from normal cells.
Lack of cytotoxicity to an HLA-A24 mutant cell line We addressed the question of whether the cytotoxicity of TAK-1 to leukemia cells was restricted by HLA-A24, as it was in the experiment on WT1 peptide-loaded target cells, by performing a conventional inhibition assay using an anti-HLA class I MoAb. As expected, the addition of an anti-HLA class I framework MoAb, w6/32, to the assay medium inhibited the cytotoxicity of TAK-1 to leukemia cell lines (Figure 3C). Furthermore, confirmation that the cytotoxicity of TAK-1 to leukemia cells was restricted by HLA-A24 was provided by the experiment using the HLA-A24 mutant cell line KU812.37 Sequence analysis of the HLA-A gene demonstrated that the 23-base repeated insertion occurred in HLA-A24 gene exon 2 (Figure 3A). This insertion resulted in a frameshift of the HLA-A24 gene sequence and the failure of HLA-A24 molecule expression (Figure 3B). As shown in Figure 3C, TAK-1 did not lyse KU812, even though KU812 expressed WT1, whereas TK91, an HLA-A24-positive myeloid leukemia cell line, was lysed by TAK-1. These data suggest strongly that the cytotoxocity of TAK-1 to leukemia cells is restricted by HLA-A24.
Reduction of TAK-1-mediated cytotoxicity to leukemia cells by treatment with a WT1 antisense oligonucleotide We carried out further investigations to establish whether TAK-1 recognizes endogenously processed WT1 peptide using an antisense oligonucleotide complementary to WT1 gene. As shown in Figure 4A, the WT1 protein expression levels of the leukemia cell lines TK91 and MEG01 were reduced considerably by treatment with an antisense, but not a random or sense oligonucleotide, complementary to the WT1 gene. Inhibition of WT1 synthesis by leukemia cells reduced the degrees of TAK-1-mediated cytotoxicity (Figure 4B). Treatment of leukemia cell lines with an antisense oligonucleotide complementary to the WT1 gene affected neither cell viability nor the expression of surface molecules important for CTL recognition, including HLA class I, ICAM-1, B7, and CD40 (data not shown). Therefore, these data indicate that TAK-1 lyses leukemia cells by recognizing the WT1 peptide in the context of the HLA-A24 molecule.
Lack of effect of TAK-1 on colony formation by normal bone marrow cells Recently, WT1 was reported to be expressed in normal bone marrow CD34+ hematopoietic progenitors, but this is still controversial.17,25,26 Therefore, we addressed the issue of whether TAK-1 recognizes WT1 peptide expressed on normal bone marrow progenitor cells and suppresses their growth. As shown in Figure 5, after coculture with TAK-1 in the absence of WT1 peptide, the numbers of CFU-GM and BFU-E generated from bone marrow cells of 2 HLA-A24-positive individuals were almost the same as those generated from bone marrow cells cultured alone. However, the numbers of CFU-GM and BFU-E decreased significantly when HLA-A24-positive bone marrow cells were pretreated with WT1 peptide and then cocultured with TAK-1. As expected, TAK-1 had no effect on colony formation by HLA-A24-negative bone marrow cells, which had been pretreated with WT1 peptide or left untreated. These data suggest strongly that WT1 is not expressed in normal bone marrow progenitors or their expression levels of WT1 are not high enough to be recognized by T lymphocytes.
Evidence that human T lymphocytes discriminate leukemia and normal
cells has been accumulating. The target molecules of
T-lymphocyte-mediated immune responses against human leukemia cells
reported most frequently in the previous papers are fusion proteins,
such as BCR-ABL,2-10 PML/RAR
We thank Drs Masafumi Takiguchi (Kumamoto University, Kumamoto, Japan),
Toshihiko Kaneshige (Shionogi Biomedical Laboratory, Osaka, Japan),
Kenji Kishi (Tokai University, Kanagawa, Japan), Masuhiro Takahashi
(Niigata University, Niigata, Japan), and Hayato Yamauchi (Ehime
University, Ehime, Japan) for providing the cell lines and for their
helpful suggestions. We also thank Kirin Brewery Co Ltd, Dainippon
Pharmaceutical Co Ltd, and Kyowa Hakko Kogyo Co Ltd, for providing
GM-CSF, TNF-
Submitted March 23, 1999; accepted September 9, 1999.
Supported by grants from the Ministry of Education, Science, Sports, and Culture of Japan; the Ministry of Health and Welfare of Japan; the Mochida Foundation for Medical and Pharmaceutical Research, Japan; the Inamori Foundation, Japan; the Suzuken Memorial Foundation, Japan; and the Naito Foundation, Japan.
Reprints: Masaki Yasukawa, the First Department of Internal Medicine, Ehime University School of Medicine, Shigenobu, Ehime 791-0295, Japan; e-mail: yasukawa{at}m.ehime-u.ac.jp.
The publication costs of this article were defrayed in part by page charge payment. Therefore, and solely to indicate this fact, this article is hereby marked "advertisement" in accordance with 18 U.S.C. section 1734.
1. Falkenburg JHF, Smit WM, Willemze R. Cytotoxic T-lymphocyte (CTL) responses against acute or chronic myeloid leukemia. Immunol Rev. 1997;157:223-230[Medline] [Order article via Infotrieve].
2.
Bocchia M, Korontsvit T, Xu Q, et al.
Specific human cellular immunity to bcr-abl oncogene-derived peptides.
Blood.
1996;87:3587-3592
3.
ten Bosch GJA, Joosten AM, Kassler JH, Melief CJM, Leeksma OC.
Recognition of BCR-ABL positive leukemic blasts by human CD4+ T cells elicited by primary in vitro immunization with a BCR-ABL breakpoint peptide.
Blood.
1996;88:3522-3527 4. Greco G, Fruci D, Accapezzato D, et al. Two bcr-abl junction peptides bind HLA-A3 molecules and allow specific induction of human cytotoxic T lymphocytes. Leukemia. 1996;10:693-699[Medline] [Order article via Infotrieve].
5.
Mannering SI, McKenzie JL, Fearnley DB, Hart DNJ.
HLA-DR1-restricted bcr-abl (b3a2)-specific CD4+ T lymphocytes respond to dendritic cells pulsed with b3a2 peptide and antigen-presenting cells exposed to b3a2 containing cell lysates.
Blood.
1997;90:290-297 6. Buzyn A, Ostankovitch M, Zerbib A, et al. Peptides derived from the whole sequence of BCR-ABL bind to several class I molecules allowing specific induction of human cytotoxic T lymphocytes. Eur J Immunol. 1997;27:2066-2072[Medline] [Order article via Infotrieve].
7.
Nieda M, Nicol A, Kikuchi A, et al.
Dendritic cells stimulate the expansion of bcr-abl specific CD8+ T cells with cytotoxic activity against leukemic cells from patients with chronic myeloid leukemia.
Blood.
1998;91:977-983
8.
Yasukawa M, Ohminami H, Kaneko S, et al.
CD4+ cytotoxic T-cell clones specific for bcr-abl b3a2 fusion peptide augment colony formation by chronic myelogenous leukemia cells in a b3a2-specific and HLA-DR-restricted manner.
Blood.
1998;92:3355-3361 9. Yotnda P, Firat H, Garcia-Pons F, et al. Cytotoxic T cell response against the chimeric p210 BCR-ABL protein in patients with chronic myelogenous leukemia. J Clin Invest. 1998;101:2290-2296[Medline] [Order article via Infotrieve]. 10. Osman Y, Takahashi M, Zheng Z, et al. Generation of bcr-abl specific cytotoxic T-lymphocytes by using dendritic cells pulsed with bcr-abl (b3a2) peptide: its applicability for donor leukocyte transfusions in marrow grafted CML patients. Leukemia. 1999;13:166-174[Medline] [Order article via Infotrieve].
11.
Gambacorti-Passerini C, Grignani F, Arienti F, Pandolfi PP, Pelicci PG, Parmiani G.
Human CD4 lymphocytes specifically recognize a peptide representing the fusion region of the hybrid protein pml/RAR 12. Yotnda P, Garcia F, Peuchmaur M, et al. Cytotoxic T cell response against the chimeric ETV6-AML1 protein in childhood acute lymphoblastic leukemia. J Clin Invest. 1998;102:455-462[Medline] [Order article via Infotrieve].
13.
Ohminami H, Yasukawa M, Kaneko S, et al.
Fas-independent and nonapoptotic cytotoxicity mediated by a human CD4+ T-cell clone directed against an acute myelogenous leukemia-associated DEK-CAN fusion peptide.
Blood.
1999;93:925-935
14.
Molldrem J, Dermime S, Parker K, et al.
Targeted T-cell therapy for human leukemia: cytotoxic T lymphocytes specific for a peptide derived from proteinase 3 preferentially lyse human myeloid leukemia cells.
Blood.
1996;88:2450-2457 15. Miwa H, Beran M, Saunders GF. Expression of the Wilms' timor gene (WT1) in human leukemias. Leukemia. 1992;6:405-409[Medline] [Order article via Infotrieve].
16.
Inoue K, Sugiyama H, Ogawa H, et al.
WT1 as a new prognostic factor and a new marker for the detection of minimal residual disease in acute leukemia.
Blood.
1994;84:3071-3079 17. Menssen HD, Renkl H-J, Rodeck U, et al. Presence of Wilms' tumor gene (wt1) transcripts and the WT1 nuclear protein in the majority of human acute leukemias. Leukemia. 1995;9:1060-1067[Medline] [Order article via Infotrieve]. 18. Call KM, Glaser T, Ito CY, et al. Isolation and characterization of a zinc finger polypeptide gene at the human chromosome 11 Wilms' tumor locus. Cell. 1990;60:509-520[Medline] [Order article via Infotrieve]. 19. Rauscher FJ III. The WT1 Wilms tumor gene product: a developmentally regulated transcription factor in the kidney that functions as a tumor suppressor. FASEB J. 1993;7:896-903[Abstract]. 20. Pritchard-Jones K, Fleming S, Davidson D, et al. The candidate Wilms' tumour gene is involved in genitourinary development. Nature. 1990;346:194-197[Medline] [Order article via Infotrieve].
21.
Huang A, Campbell CE, Bonetta L, et al.
Tissue, developmental, and tumor-specific expression of divergent transcripts in Wilms tumor.
Science.
1990;250:991-994 22. Kreidberg JA, Sariola H, Loring JM, et al. WT-1 is required for early kidney development. Cell. 1993;74:679-691[Medline] [Order article via Infotrieve]. 23. Park S, Schalling M, Bernard A, et al. The Wilms tumour gene WT1 is expressed in murine mesoderm-derived tissues and mutated in a human mesothelioma. Nat Genet. 1993;4:415-420[Medline] [Order article via Infotrieve].
24.
Pelletier J, Schalling M, Buckler AJ, Rogers A, Haber DA, Housman DE.
Expression of the Wilms' tumor gene WT1 in the murine urogenital system.
Genes Dev.
1991;5:1345-1356
25.
Inoue K, Ogawa H, Sonoda Y, et al.
Aberrant overexpression of the Wilms tumor gene (WT1) in human leukemia.
Blood.
1997;89:1405-1412 26. Maurer U, Brieger J, Weidmann E, Mitrou PS, Hoelzer D, Bergmann L. The Wilms' tumor gene is expressed in a subset of CD34+ progenitors and downregulated early in the course of differentiation in vitro. Exp Hematol. 1997;25:945-950[Medline] [Order article via Infotrieve].
27.
Inoue K, Ogawa H, Yamagami T, et al.
Long-term follow-up of minimal residual disease in leukemia patients by monitoring WT1 (Wilms tumor gene) expression levels.
Blood
1996;88:2267-2278
28.
Bergmann L, Miething C, Maurer U, et al.
High levels of Wilms' tumor gene (wt1) mRNA in acute myeloid leukemias are associated with a worse long-term outcome.
Blood.
1997;90:1217-1225 29. Karaki S, Kariyone A, Kato N, Iwakura Y, Takiguchi M. HLA-B51 transgenic mice as recipients for production of polymorphic HLA-A, B-specific antibodies. Immunogenetics. 1993;37:139-142[Medline] [Order article via Infotrieve].
30.
Tanaka F, Fujie T, Tahara K, et al.
Induction of antitumor cytotoxic T lymphocytes with a MAGE-3-encoded synthetic peptide presented by human leukocytes antigen-A24.
Cancer Res.
1997;57:4465-4468 31. Ibe M, Moore YI, Miwa K, Kaneko Y, Yokota S, Takiguchi M. Role of strong anchor residues in the effective binding of 10-mer and 11-mer peptides to HLA-A*2402 molecules. Immunogenetics. 1996;44:233-241[Medline] [Order article via Infotrieve]. 32. Tokunaga K, Ishikawa Y, Ogawa A, et al. Sequence-based association analysis of HLA class I and II alleles in Japanese supports conservation of common haplotypes. Immunogenetics. 1997;46:199-205[Medline] [Order article via Infotrieve]. 33. Kaneshige T, Hashimoto M, Matsumoto Y, et al. Serologic and nucleotide sequencing analyses of a novel DR52-associated DRB1 allele with the DR `NJ25' specificity, designated DRB1*1307. Hum Immunol. 1994;41:151-159[Medline] [Order article via Infotrieve]. 34. Yasukawa M, Inatsuki A, Horiuchi T, Kobayashi Y. Functional heterogeneity among herpes simplex virus-specific human CD4+ T cells. J Immunol. 1991;146:1341-1347[Abstract].
35.
Yasukawa M, Yakushijin Y, Hasegawa H, et al.
Expression of perforin and membrane-bound lymphotoxin (tumor necrosis factor-
36.
Yamagami T, Sugiyama H, Inoue K, et al.
Growth inhibition of human leukemic cells by WT1 (Wilms tumor gene) antisense oligodeoxynucleotides: implications for the involvement of WT1 in leukemogenesis.
Blood.
1996;87:2878-2884 37. Kishi K. A new leukemia cell line with Philadelphia chromosome characterized as basophil precursors. Leuk Res. 1985;9:381-390[Medline] [Order article via I |
Top
Abstract
Introduction
1, insulin-like growth factor II, and its
own gene promoters.
